RESUMEN
Oxidative stress triggered by aging, radiation, or inflammation impairs ovarian function by inducing granulosa cell (GC) apoptosis. However, the mechanism inducing GC apoptosis has not been characterized. Here, we found that ovarian GCs from aging patients showed increased oxidative stress, enhanced reactive oxygen species activity, and significantly decreased expression of the known antiapoptotic factor sphingosine-1-phosphate/sphingosine kinase 1 (SPHK1) in GCs. Interestingly, the expression of Krüppel-like factor 12 (KLF12) was significantly increased in the ovarian GCs of aging patients. Furthermore, we determined that KLF12 was significantly upregulated in hydrogen peroxide-treated GCs and a 3-nitropropionic acid-induced in vivo model of ovarian oxidative stress. This phenotype was further confirmed to result from inhibition of SPHK1 by KLF12. Interestingly, when endogenous KLF12 was knocked down, it rescued oxidative stress-induced apoptosis. Meanwhile, supplementation with SPHK1 partially reversed oxidative stress-induced apoptosis. However, this function was lost in SPHK1 with deletion of the binding region to the KLF12 promoter. SPHK1 reversed apoptosis caused by hydrogen peroxide-KLF12 overexpression, a result further confirmed in an in vitro ovarian culture model and an in vivo 3-nitropropionic acid-induced ovarian oxidative stress model. Overall, our study reveals that KLF12 is involved in regulating apoptosis induced by oxidative stress in aging ovarian GCs and that sphingosine-1-phosphate/SPHK1 can rescue GC apoptosis by interacting with KLF12 in negative feedback.
Asunto(s)
Envejecimiento , Apoptosis , Células de la Granulosa , Peróxido de Hidrógeno , Factores de Transcripción de Tipo Kruppel , Lisofosfolípidos , Fosfotransferasas (Aceptor de Grupo Alcohol) , Esfingosina , Femenino , Humanos , Envejecimiento/metabolismo , Retroalimentación Fisiológica , Células de la Granulosa/efectos de los fármacos , Células de la Granulosa/metabolismo , Peróxido de Hidrógeno/farmacología , Técnicas In Vitro , Factores de Transcripción de Tipo Kruppel/antagonistas & inhibidores , Factores de Transcripción de Tipo Kruppel/biosíntesis , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Lisofosfolípidos/biosíntesis , Lisofosfolípidos/metabolismo , Técnicas de Cultivo de Órganos , Estrés Oxidativo/efectos de los fármacos , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Regiones Promotoras Genéticas , Esfingosina/biosíntesis , Esfingosina/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Meiosis-arrest female 1 (MARF1) is a recently identified key oogenic regulator essential for the maintenance of female fertility and genome integrity in mice. However, the detailed functions and the underlying mechanisms of MARF1 remain elusive. Here, in an attempt to create a mouse model expressing fluorescent protein-tagged MARF1 to facilitate further exploration of the roles of MARF1 in oocytes, we produced a Marf1-eGFP knockin (KI) mouse line in which the C-terminal structure and function of MARF1 were interfered by its fusing eGFP peptide. Using these Marf1-eGFP-KI mice, we revealed, unexpectedly, the functions of MARF1 in the control of oocyte meiotic division. We found that the Marf1-eGFP-KI females ovulated mature oocytes with severe meiotic and developmental defects, and thus were infertile. Moreover, meiotic reinitiation was delayed while meiotic completion was accelerated in the KI-oocytes, which was coincident with the increased incidence of oocyte aneuploidy. Therefore, MARF1 is indispensable for maintaining the fidelity of homolog segregation during oocyte maturation, and this function relies on its C-terminal domains.
RESUMEN
Hypoxia-inducible factor-1 (HIF-1) orchestrates angiogenesis under hypoxic conditions mainly due to increased expression of such target genes as vascular endothelial growth factor (VEGF). Na+/H+exchanger-1 (NHE1), a potential HIF target gene product, plays a pivotal role in proliferation, survival, migration, adhesion and so on. However, it is unknown whether NHE1 is involved in HIF-1α-induced angiogenesis. This present study demonstrated that the expression of NHE1 was much higher in human umbilical vein endothelial cells (HUVECs) infected with adenovirus encoding HIF-1α (rAd-HIF) than with vacuum adenovirus (vAd). HIF-1α also increased the expression of VEGF, the expression and activity of calpains, and the intracellular pH. Moreover, small interfering RNA targeting NHE1 (NHE1 siRNA) dramatically decreased the expression of NHE1 and thus lowered the intracellular pH, and it also attenuated the protein expression of calpain-2 but not calpain-1, resulting in the lower calpain activity. Furthermore, HIF-1α enhanced the proliferation, migration and Matrigel tube formation, which were inhibited by NHE1 siRNA. Finally, the inhibitory effect of NHE1 siRNA was reversed by VEGF and the reversibility of the later was abrogated by the calpain inhibitor ALLM. In conclusion, the findings have revealed that NHE1 might participate in HIF-1-induced angiogenesis due, at least in part, to the alteration of the calpain activity, suggesting that NHE1 as well as calpains might represent a potential target of controlling angiogenesis in response to the hypoxic stress under various pathological conditions.
Asunto(s)
Calpaína/metabolismo , Proteínas de Transporte de Catión/deficiencia , Células Endoteliales/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Neovascularización Fisiológica/fisiología , ARN Interferente Pequeño/genética , Calpaína/antagonistas & inhibidores , Proteínas de Transporte de Catión/genética , Proteínas de Transporte de Catión/metabolismo , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/genética , Supervivencia Celular/fisiología , Células Cultivadas , Inhibidores de Cisteína Proteinasa/farmacología , Citoplasma/metabolismo , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Expresión Génica/genética , Humanos , Concentración de Iones de Hidrógeno , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Neovascularización Fisiológica/efectos de los fármacos , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno/genética , Intercambiadores de Sodio-Hidrógeno/metabolismo , Transducción Genética , Regulación hacia Arriba/genética , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor A de Crecimiento Endotelial Vascular/farmacologíaRESUMEN
The kinds and the distribution of the coal in China are investigated. The results indicated that the 80% coal in China is used by the method of the coal gasification. The possibility of utilization and development of the fuel cell power plant in China is analyzed. A combined cycle generation system is designed. Its net electrical efficiency is about 55% (LHV), which is higher than that of the fire power plant. So it is environmental-friendly and high-efficient generation mode.